Human cytomegalovirus (HCMV), a herpesvirus, is a major opportunistic infectious agent in individuals suffering from AIDS, as well as individuals with suppressed immune systems (for example, organ and bone marrow transplant recipients). Both primary and reactivated latent HCMV infections can cause severe acute diseases in these individuals, such as retinitis, pneumonitis, hepatitis and gastroenteritis. Current therapies against HCMV infections are mostly targeted at the DNA polymerase of the virus, and are limited in their usefulness by their toxic side effects. In addition, viral resistance to anti-herpes agents is becoming an increasingly more significant problem. Therefore, new and efficacious treatments for HCMV infections, and herpesvirus infections in general, are highly desirable. The protease of herpesviruses is essential for their life cycle, and represents a novel target for the design and development of anti-herpes chemotherapeutic agents. Several classes of inhibitors against HCMV protease have been reported, but none of these have sufficient potency and/or pharmacokinetic properties. Breakthroughs are needed to develop a new generation of inhibitors against the protease, with higher potency, metabolic stability, and oral bioavailability. Structure-based drug design can play an important role in this process, as it has in the development of AIDS therapeutic agents targeted at the HIV protease. Such design efforts require a detailed structural and biochemical knowledge of the protein target, which are currently still lacking for HCMV protease. Despite being a serine protease, HCMV protease has many unique biochemical and structural features and belongs to a new class of serine protease, distinct from the classical serine proteases such as chymotrypsin and subtilisin. Therefore, a new body of knowledge is needed on this new class of enzymes. The proposed research will use structural, biochemical and biophysical techniques to achieve a greater understanding of the molecular basis for the inhibition and the catalytic mechanism of HCMV protease. Special emphasis will be placed on studying the unique features of the protease, such as the Ser-His-His catalytic triad, the requirement for dimerization for activity, the activation by antichaotropic agents, the conformational flexibility and the induced fit behavior, and the inhibition of the protease by non-peptidic and peptidomimetic compounds. The crystal structure of HCMV protease free enzyme and the recently determined structure of the protease inhibitor complex represent an excellent starting point for the performance and completion of the proposed research. A long-term goal of the research is to expand the studies to include the proteases of other herpesviruses, many of which (herpes simplex virus and Kaposi's sarcoma associated herpesvirus) are also targets for the development of anti-herpes agents.